The importance of alkaline phosphatase (orthophosphoric-monoester phosphohydrolase EC 3.1.3.1) in clinical medicine and molecular biology has made it a popular subject for scientific study and commercial utility. (See for example, H. N. Ferley, "The Enzymes," Academic Press, New York Vol. IV, pp. 417-447, (1971); R. B. McComb, et al., "Alkaline Phosphatase," Plenum Press, New York (1979); B. L. Vallee and D. S. Auld, Biochem. 32:6494-6500 (1993)).
Alkaline phosphatase has been purified and characterized from a variety of bacterial, fungal, alga, invertebrate and vertebrate species (R. B. McComb et al., supra). The enzyme also has been purified from mesophiles and thermophiles. A relatively unstable alkaline phosphatase was characterized from a thermophilic Thermus species. (A. T. Hartog et al. Int. J. Biochem., 24, 1657-1660 (1992), but alkaline phosphatase has not, until now, been purified and characterized from a hyperthermophile.
Hyperthermophiles are a group of microorganisms which grow at 80.degree. C. or above. Extremely thermostable enzymes produced by these organisms have attracted increased attention because they allow structural and functional studies of proteins at very high temperatures and have many molecular biology-related, and potential industrial, applications. (See for example, M. W. W. Adams Annu. Rev. Microbiol. 47:627-658 (1993)); T. Coolbear et al. Adv. Biochem. Eng./Biotechol. 45:57-98 (1992)). The enzymes from hyperthermophilic microorganisms also provide opportunities to investigate the structure and function of proteins at very high temperatures.
Thermotoga is a hyperthermophilic eubacterium that grows optimally at 80.degree. C. and is the most thermophilic eubacterium discovered to date. (H. W. Jannasch et al., Arch Microbiol. 150:103-104, (1988)).
The present invention provides alkaline phosphatase from Thermatoga neapolitana in purified form and a method of effecting that purification.